Rapid identification of candidate genes controlling male-sterility in Foxtail millet (Setaria italica).

Photo-(thermo-) sensitive genic male-sterile line is the key component of two-line hybridization system in foxtail millet (Setaria italica), but the genetic basis of male sterility in most male-sterile lines is still unclear. In the present study, a large F2 population was developed derived from a cross between the photo-(thermo-) sensitive male-sterile line A2 and the fertile-line 1484-5. Thirty plants with extreme high and extreme low fertility were selected from the population to construct a sterile DNA pool and a fertile DNA pool, respectively. Sequencing both DNA pools and data analysis revealed that two QTLs conferred male-sterility, qSiMS6.1 with a major effect and qSiMS6.2 with a minor effect, on chromosome 6. Both QTLs exhibited complete dominance. The major QTL, qSiMS6.1, was delimited to a 186-kb interval between the markers SiM20 and SiM9 by the joint analysis of QTL-seq and QTL mapping with SSR and structure variation markers. Millet_GLEAN_10020454 in this region is the most likely candidate gene for qSiMS6.1 since it is predicted to encode a male-sterile 5 like protein. These results lay a solid foundation for qSiMS6.1 cloning and provided gene resources for breeding new male-sterile lines. Supplementary Information: The online version contains supplementary material available at 10.1007/s11032-021-01269-2.

QTL analysis of important agronomic traits and metabolites in foxtail millet (Setaria italica) by RIL population and widely targeted metabolome.

As a bridge between genome and phenotype, metabolome is closely related to plant growth and development. However, the research on the combination of genome, metabolome and multiple agronomic traits in foxtail millet (Setaria italica) is insufficient. Here, based on the linkage analysis of 3,452 metabolites via with high-quality genetic linkage maps, we detected a total of 1,049 metabolic quantitative trait loci (mQTLs) distributed in 11 hotspots, and 28 metabolite-related candidate genes were mined from 14 mQTLs. In addition, 136 single-environment phenotypic QTL (pQTLs) related to 63 phenotypes were identified by linkage analysis, and there were 12 hotspots on these pQTLs. We futher dissected 39 candidate genes related to agronomic traits through metabolite-phenotype correlation and gene function analysis, including Sd1 semidwarf gene, which can affect plant height by regulating GA synthesis. Combined correlation network and QTL analysis, we found that flavonoid-lignin pathway maybe closely related to plant architecture and yield in foxtail millet. For example, the correlation coefficient between apigenin 7-rutinoside and stem diameter reached 0.98, and they were co-located at 41.33-44.15 Mb of chromosome 5, further gene function analysis revealed that 5 flavonoid pathway genes, as well as Sd1, were located in this interval . Therefore, the correlation and co-localization between flavonoid-lignins and plant architecture may be due to the close linkage of their regulatory genes in millet. Besides, we also found that a combination of genomic and metabolomic for BLUP analysis can better predict plant agronomic traits than genomic or metabolomic data, independently. In conclusion, the combined analysis of mQTL and pQTL in millet have linked genetic, metabolic and agronomic traits, and is of great significance for metabolite-related molecular assisted breeding.

Metabolome-Based Genome-Wide Association Study Provides Genetic Insights Into the Natural Variation of Foxtail Millet.

The plant metabolome is considered as a bridge between the genome and the phenome and is essential for the interaction between plant growth and the plant environment. Here, we used the liquid chromatography-tandem mass spectrometry method to perform a widely targeted metabolomics analysis of 150 millet germplasm and simultaneous identification and quantification of 330 annotated metabolites. Comparing the metabolic content of different millets revealed significant natural variation of both primary and secondary metabolites, including flavonoids, phenolamides, hydroxycinnamoyl derivatives, nucleotides, and lipids, in the millets from India and the north and south of China; among them, some of the flavonoids are the most prominent. A total of 2.2 TB sequence data were obtained by sequencing 150 accessions of foxtail millet using the Illumina platform. Further digging into the genetic basis of metabolites by mGWAS analysis found that cyanidin 3-O-glucoside and quercetin O-acetylhexside are concentratedly located at 43.55 Mb on chromosome 5 and 26.9 Mb on chromosome 7, and two Lc were mined as candidate genes, respectively. However, the signals of luteolin 7-O-glucoside and kaempferol 3-O-glucoside were also detected at 14.36 Mb on chromosome 3, and five glycosyltransferase genes on this loci were deemed to regulate their content. Our work is the first research to use mGWAS in millet, and it paves the way for future dissection of complex physiological traits in millet.

Comprehensive Profiling and Inheritance Patterns of Metabolites in Foxtail Millet.

Metabolomics aims at determining a sample's metabolites profile and hence provides a straight functional statement of an organism's physiological condition. Here, we investigated comprehensive profiling, natural variation and species-specific accumulation of both primary and secondary metabolites in foxtail millet using LC-MS, and inheritance patterns of metabolome in millet hybrids. The application of a broad target metabolomics method facilitated the simultaneous identification and quantification of more than 300 metabolites. The metabolic analysis of these compounds, such as flavonoids, phenolamides, hydrocinnamoyl derivatives, vitamins and LPCs, revealed their developmentally controlled accumulation, and natural variation in different tissues/varieties. Species-specific accumulation of secondary metabolites was observed based on a comparative metabolic analysis between millet and rice, such as flavonoid O-rutinosides/neohesperidosides and malonylated flavonoid O-glycosides. In analyzing the metabolic variations between hybrid progenies and their parental lines, including a photothermo-sensitive genic male sterility line and five Zhangzagu varieties, metabolic overdominant, and dominant patterns of inheritance could be observed. For example, hydrocinnamoyl derivatives and feruloylated flavonoids were identified as over-parent heterosis (overdominant) metabolites in milet hybrids. Our work paves the way for developing predictors of hybrid performance and the future analysis of the biosynthesis and regulation of relevant metabolic pathways in millet.

Identification of QTLs for 14 Agronomically Important Traits in Setaria italica Based on SNPs Generated from High-Throughput Sequencing.

Foxtail millet (Setaria italica) is an important crop possessing C4 photosynthesis capability. The S. italica genome was de novo sequenced in 2012, but the sequence lacked high-density genetic maps with agronomic and yield trait linkages. In the present study, we resequenced a foxtail millet population of 439 recombinant inbred lines (RILs) and developed high-resolution bin map and high-density SNP markers, which could provide an effective approach for gene identification. A total of 59 QTL for 14 agronomic traits in plants grown under long- and short-day photoperiods were identified. The phenotypic variation explained ranged from 4.9 to 43.94%. In addition, we suggested that there may be segregation distortion on chromosome 6 that is significantly distorted toward Zhang gu. The newly identified QTL will provide a platform for sequence-based research on the S. italica genome, and for molecular marker-assisted breeding.